A major aim is to continue studies on the in vivo roles of eukaryotic DNA topoisomerases using yeast as a model system. Two new yeast topoisomerases, products of the TOP3 and UPRI genes, will be investigated, as will the functional overlap of these enzymes with the previously known DNA topoisomerase I. Suppresor genes that, when mutated, improve the growth of various topoisomerase mutants, will be cloned and analyzed. The gene(s) coding for the two forms of Xenopus DNA topoisomerase I will be cloned and used to study the developmentally regulated expression of these enzymes. A novel genetic system will be employed to study interactions among the various proteins involved in transcriptional repression of the yeast silent mating-type loci. The chromatin at these genes may be similar to heterochromatin in higher eukaryotes. Also, a new approach for finding yeast DNA replication genes will be employed. Finally, the gene for a yeast cruciform-cutting endonuclease will be investigated. This enzyme may be involved in resolving the branched recombination intermediate known as the Holliday junction. Therefore, the effect of mutations in this gene on recombination will be studied.